Abstract Chronic exposure to ethanol (EtOH) has a variety of adverse effects on health, and is responsible for a substantial amount of morbidity and mortality in the US and worldwide. Evidence that exposure to EtOH in parents can have effects on their offspring has raised the possibility that EtOH has effects on the epigenome. The epigenome is the complex assortment of molecular structures that are associated with the genome to control gene expression and cell differentiation. An epigenetic variant is a change in the molecular components of the epigenome at a specific transcriptional regulatory element and in a specific cell type: it is functionally relevant if it results in a stable change in the expression of a gene. Epigenetic variants induced in the germline can be passed to succeeding generations, a phenomenon termed epigenetic inheritance. The extent to which epigenetic variants can be induced by EtOH, and transmitted from parents to offspring through multiple generations, has never been established. We hypothesize that paternal preconception EtOH exposure induces epigenetic variants that are heritable in the absence of any genetic variation. Since changes in gene expression levels are the functional outcome of epigenetic variants, we will use gene expression variants, in a single homogeneous cell type, as the functional readout of epigenetic variation; the study will be carried out in an isogenic mouse strain in order to minimize confounding genetic and environmental factors, and use an established system of ethanol exposure. We will ask if paternal preconception EtOH exposure can induce epigenetic variants that persist for a generation past the cessation of EtOH exposure. The study will assess the frequency with which paternal preconception EtOH exposure induces gene expression variants in the offspring of exposed males, and the frequency with which such variants are transmitted to the next generation. This proposal addresses directly the subject of PA-13-004, Epigenetic Inheritance and Transgenerational Effects of Alcohol; evidence for the heritability of EtOH-induced epigenetic variants would have major public health implications.